Full length articleHost macrophage response to injectable hydrogels derived from ECM and α-helical peptides
Graphical abstract
Introduction
The repair or replacement of damaged tissue using tissue engineering strategies is strongly influenced by the material selected to mimic the architecture and functional characteristics of native healthy tissue. Published sources of bespoke materials are either biologically derived or synthetic [1,2]. In vitro response to these biomaterials is usually assessed by their ability to promote specific cell responses. However, it is the host immune response to implanted biomaterials that is the critical, if not the defining, determinant of clinical success or failure [3,4]. A desirable host response relies on cell infiltration and material integration/ remodelling in support of an optimal functional outcome. Prolonged inflammation typically results in the formation of granulation tissue or fibrous capsules [5], seroma, scars and encapsulation [6]. Such an effect results in the isolation of the implanted material from the surrounding healthy tissue and prevents the formation of new functional tissue. Macrophages represent a major cellular component of the innate immune response to biomaterials. These cells show diverse plasticity in their functions ranging from pro-inflammatory to anti-inflammatory and reparative phenotypes [7,8]. Contrary to accepted 25-year dogma, macrophages, among other cell types such as muscle-specific regulatory T cells and satellite cells [9], are essential for normal tissue development [10], [11], [12], [13]. Macrophages are necessary for successful tissue and organ regeneration in regenerative species such as the axolotl [14,15], and have the ability to affect stem cell/ progenitor cell differentiation [16], and proliferation (i.e., are not necessarily “end stage” cells) [17]. Given this relatively recent understanding of macrophage biology and their role in critical life processes, the signalling molecules, physical factors, and environmental factors that influence macrophage phenotype are of great interest to the biomaterials community.
Naturally occurring biomaterials composed of extracellular matrix (ECM), such as decellularised tissues [18], [19], [20], have been shown to contain a variety of potent signalling molecules that are released or exposed during degradation of the matrix. These include cryptic peptides [14], cytokines and chemokines [20,21], and, most recently, embedded matrix bound nanovesicles (MBV) [22]. In addition to the chemical cues, these materials provide physical signals such as material stiffness, pore size, and load transfer [20], which have all been shown to influence macrophage phenotype [23]. Several tissue types have been decellularised and used either in their original form, as sheets [24], or as injectable hydrogels [20]. The tissues used include dermis [25,26], small intestinal submucosa (SIS) [27], [28], [29], urinary bladder matrix (UBM) [30,31], liver [32], tendons [33], and whole limbs [34]. The use of decellularised ECM in clinical applications is commonplace with several FDA-approved ECM products currently available on the market including AlloDermⓇ, NeoForm™, GraftJacketⓇ, Strattice™, Meso BioMatrixⓇ, SynerGraftⓇ OasisⓇ and SurgisisⓇ.
There have been many attempts to mimic properties of the native ECM within synthetic biomaterials [35], [36], [37]. It is possible to study individual ECM proteins and to design peptides that are capable of hydrogel configurations [38], [39], [40], [41], [42]. Some aspects of the complexity of naturally occurring ECM can be incorporated through the addition of cell-guiding chemical moieties [43], [44], [45]. The use of synthetic peptides to accomplish these ends is not new, and there are several reports of natural proteins that are used to create hydrogels suitable for clinical applications [46], [47], [48]. The benefits of the previously described hydrogelating self-assembling fibre (hSAF) system include their stable α-helical structure, modifiable chemistry and ability to form at low concentrations compared to other peptide-based systems [42,43,49]. Briefly, the hSAF system is formed from two complementary de novo designed α-helical peptides. The two peptide sequences form sticky-ended dimers that assemble end-to-end to form fibres. At mM concentrations, the mixed hSAF peptide fibres form self-supporting hydrogels. Combined with added chemical functionality, such as the addition of adhesive ligands, these gels offer at least partial control over cell adhesion, migration and differentiation [43], all characteristics of a material with high clinical translational potential.
While there are clear compositional differences between SIS, UBM and hSAFs, they all form hydrogels with fibrous networks through which cells can migrate and form three-dimensional constructs. Herein, we investigate the macrophage response to injectable UBM, SIS, bovine collagen (type I; FibriCol) and hSAF hydrogels in a partial thickness abdominal wall defect model in Sprague Dawley rats over 28 days, using the native tissue for comparison. Macrophage interaction with these materials will provide a pre-clinical indicator of the likely cell response in vivo and offer insights into strategies for promoting cell infiltration and functional restoration of damaged tissue.
Section snippets
Small Intestinal Submucosa
Porcine small intestinal submucosa (SIS) was prepared using previously described methods [50]. Briefly, the intestine was flushed with double distilled water, opened horizontally and most of the tunica mucosa, and entirety of tunica serosa and tunica muscularis externa were removed using a scraper. The remaining tunica submucosa and basilar layers of the tunica mucosa were cut to 80 g particles and shaken at 300 rpm for 2 hrs at room temperature in 0.1% peracetic acid (v/v) (Rochester Midland,
In Vitro Effect of ECM-derived and α-Helical Peptide-based Hydrogels on Primary Murine Macrophages
In vitro, the presence of the hydrogels did not inhibit monocyte differentiation into macrophages. As shown in Figs. 2A and 2B, at least 50% of the cells seeded onto the hSAF, collagen, SIS, and UBM hydrogels were positive for the pan-macrophage marker F4/80, indicating successful differentiation, without significant differences between the different hydrogel types.
In addition to supporting the differentiation of monocytes into macrophages, the presence of the hydrogels induced a spontaneous
Discussion
Biomaterials that attempt to mimic the chemical and physical complexity of native tissues provide controllable environments that may actively promote cell attachment and differentiation [56], [57], [58], [59]. Hypothetically, these materials are more likely to generate a constructive and functional response in vivo, lead to integration with host tissue and minimise the risk of adverse immunological responses. We investigate the macrophage response to complex injectable hydrogels derived from
Conclusion
Hydrogels derived from decellularised extracellular matrix and synthesised de novo designed α-helical peptides have been injected into rat partial thickness abdominal wall defects to examine the immune macrophage response in vivo. Over 28 days a progressive increase in mononuclear cell infiltration is observed with no foreign body reaction at the hydrogel-tissue interface, no evidence of hydrogel encapsulation or formation of multinucleate giant cells. Furthermore, the upregulation of myogenic
Declaration of Competing Interest
The authors declare no conflict of interest.
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Data Availability
The raw/processed data required to reproduce these findings cannot be shared at this time due to technical or time limitations.
Acknowledgments
This research was supported by The Royal Free Charity (Royal Free Charity Fund 190). N.M. was supported by the Engineering and Physical Sciences Research Council (EP/R02961X/1). D.A.S was supported by the BBSRC South West Biosciences Doctoral Training Partnership (BB/J014400/1 and BB/M009122/1). D.N.W. held a Royal Society Wolfson Research Merit Award (WM140008). The funders had no role in study design; in the collection, analysis and interpretation of data; in the writing of the report; and in
CRediT Author Statement
Nazia Mehrban: Conceptualization; Data curation; Formal analysis; Investigation; Methodology; Project administration; Validation; Visualization; Roles/Writing - original draft; Writing - review & editing. Catalina Pineda Molina: Conceptualization; Data curation; Formal analysis; Investigation; Methodology; Project administration; Validation; Visualization; Roles/Writing - original draft; Writing - review & editing. Lina M. Quijano: Data curation; Investigation; Methodology; Validation;
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